Regulation of the mouse cartilage-derived retinoic acid-sensitive protein gene by the transcription factor AP-2.
The expression of cartilage-derived retinoic acid-sensitive protein (CD-RAP) is initiated at the beginning of chondrogenesis and continues throughout the cartilage development. In chondrocytes, CD-RAP is down-regulated by retinoic acid. To understand the molecular mechanism underlying this regulation and the cell-specific expression, the deletion constructs of the mouse CD-RAP promoter were transfected into chondrocytes and a melanoma cell line. The results revealed a domain that demonstrated high levels of expression specifically in chondrocytes. In this functional domain, we show that a cis-acting element, 5'-GCCTGAGGC-3', binds to the trans-acting factor protein AP-2. Mutation of the AP-2 site on the CD-RAP promoter led to decreased transcription in C5.18 chondrocytes, indicating that this site may act as an activator of transcription. In contrast, increased concentration of AP-2, stimulated by retinoic acid, led to decreased transcription of the CD-RAP promoter, an effect that was abolished by mutation of the AP-2 binding site. The effect of AP-2 was further examined by co-transfection of C5.18 and HepG2 cells with the CD-RAP promoter constructs and an AP-2 expression plasmid. In a dose-dependent manner, cotransfection with AP-2 elevated and then decreased CD-RAP promoter activity. Taken together, these results suggest that AP-2 is involved in the biphasic regulation of CD-RAP transcription.
Xie
W F
WF
Department of Orthopaedics, University of Washington and Veterans Administration Puget Sound Health Care Systems, Seattle, Washington 98108, USA.
Kondo
S
S
Sandell
L J
LJ
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D002356:Cartilage / Q000378:metabolism*; D019902:Chondrocytes / Q000378:metabolism; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D016326:Extracellular Matrix Proteins; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D008546:Melanoma, Experimental / Q000378:metabolism; D051379:Mice; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics; D017384:Sequence Deletion; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014212:Tretinoin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D016326:Extracellular Matrix Proteins; C513263:Mia protein, mouse; D011506:Proteins; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D014212:Tretinoin